Conotruncal defects are clinically serious congenital heart malformations affecting the outflow tract of the developing heart. The etiologies of these conotruncal heart defects are largely unknown, but they are likely heterogeneous, involving environmental and genetic factors. The proposed research program will continue our focus on detecting genetic contributions to conotruncal heart defects. We have been supported by NHLBI R01 HL092330 to perform array comparative hybridization of 391 infants with conotruncal defects. We found that nearly 2% of boys with tetralogy of Fallot have an extra sex chromosome material. We have also successfully employed bacterial artificial chromosome array comparative genomic hybridization (array-CGH) to detect seven infants with novel submicroscopic chromosomal deletions involving chromosomes 22q11, 1q, and 10q23p and duplications of 8q21 and 8p23. Based on a novel microdeletion of the gene CRKL that we found by array-CGH, we will perform Next Generation DNA sequencing to identify mutations of genes of the signaling pathway FAK-CRKL-Erk1/Erk2 among 851 California infants with conotruncal heart defects. Murine experiments have shown that this pathway is an essential signaling pathway for cranial neural crest cells that migrate into the developing outflow tract of the heart. But there is almost nothing known about the potential contribution of sequence variation of the genes in this pathway to conotruncal defects in humans. We will also investigate three chromosomal regions where we found deletions and a duplication using array-CGH. We will use association studies of the three loci to narrow the candidate chromosomal regions. Then, we will turn to e Next Generation DNA sequencing again to identify mutations of candidate genes or regulatory elements that are located within the chromosomal microdeletions/duplication. The results of this research should lead to the development of more comprehensive, clinically applicable testing for children born with conotruncal heart defects. Our study populations of infants with conotruncal defects are unusually large and offer unique population-based investigations of children born with conotruncal defects. Overall, our research program attempts to enhance our scientific understanding of the genetic causes of conotruncal defects. Because conotruncal defects result in substantial morbidity, as well as high emotional and economic costs, expanding our understanding of their causes may lead to preventive interventions that would greatly benefit public health and society. PUBLIC HEALTH RELEVANCE: Conotruncal heart defects are cyanotic, life-threatening birth defects that are very costly to society and that require medical interventions through childhood and adulthood. Infants with conotruncal heart defects require major heart surgery during infancy, and frequently repeated surgical interventions. Almost nothing is known about the causes of conotruncal heart defects, but the frequency of familial recurrences strongly suggests there are genetic contributions. The objective of our research is identify submicroscopic chromosomal abnormalities that cause conotruncal heart defects, with a goal that improved understanding of causes may lead to better clinical testing to identify children with chromosomal microdeletions, which would then allow individualized preventive care for those children, allow more informed reproductive planning for their parents and the children, an hopefully, development of improved preventive strategies.